Methods and compositions for preparing cardiomyocytes from stem cells and uses thereof

Abstract

The present invention discloses novel compositions and methods for enhancing cardiac differentiation efficiency of stem cells or promoting ventricular and atrial cardiomyocytes formation from stem cells. The present invention also discloses the atrial and ventricular cardiomyocytes formed from the stem cells, and the uses of the cardiomyocytes for repairing cardiac injuries and screening for new medicaments for treating cardiac injuries.

What is claimed is: 
     
       1. A composition comprising a mesodermal cell prepared in vitro and an exogenous agent that inhibits the retinoic acid signaling pathway in the mesodermal cell, wherein the retinoic acid signaling pathway of the mesodermal cell is inhibited by the exogenous agent thereby promoting ventricular cardiomyocyte formation from the mesodermal cell. 
     
     
       2. The composition of  claim 1 , wherein the mesodermal cell is differentiated in vitro from a pluripotent stem cell, a totipotent stem cell, a multipotent stem cell, an oligopotent stem cell, or a unipotent stem cell. 
     
     
       3. The composition of  claim 1 , wherein the mesodermal cell is differentiated in vitro from a mammalian stem cell. 
     
     
       4. The composition of  claim 1 , wherein the mesodermal cell is differentiated in vitro from a human stem cell. 
     
     
       5. The composition of  claim 1 , wherein the mesodermal cell is prepared in vitro by contacting a stem cell with at least one of basic fibroblast growth factor (bFGF), BMP 4 and activin A. 
     
     
       6. The composition of  claim 1 , wherein the mesodermal cell is prepared in vitro by contacting a stem cell with Wnt-3a, Bio, or CHIR99021. 
     
     
       7. The composition of  claim 1 , wherein the exogenous agent is selected from the group consisting of a retinoic acid antagonist, a retinoic acid receptor antagonist, and a retinoic X receptor antagonist. 
     
     
       8. The composition of  claim 1 , wherein the retinoic acid signaling pathway of the mesodermal cell is inhibited by contacting the mesodermal cell with the exogenous agent that reduces vitamin A, or by contacting the mesodermal cell with the exogenous agent that does not comprise vitamin A. 
     
     
       9. A cell culture medium, which comprises a population of cells differentiated in vitro from a mesodermal cell whose retinoic acid signaling pathway has been inhibited by an exogenous agent, wherein, without enrichment or isolation, at least 60% of the cells in the population are cardiomyocytes, and wherein at least 80% of the cardiomyocytes are ventricular cardiomyocytes. 
     
     
       10. The cell culture medium of  claim 9 , wherein at least 90%, at least 95%, at least 99%, or 100% of the cardiomyocytes in the cell population are ventricular cardiomyocytes. 
     
     
       11. The composition of  claim 1 , wherein the mesodermal cell is differentiated in vitro from an embryonic stem cell, an induced pluripotent stem cell, a fetal stem cell, or an adult stem cell. 
     
     
       12. The composition of  claim 1 , wherein the mesodermal cell is differentiated in vitro from a human embryonic stem cell or a human induced pluripotent stem cell. 
     
     
       13. The composition of  claim 1 , wherein the exogenous agent is a pan-retinoic acid receptor antagonist. 
     
     
       14. The composition of  claim 1 , wherein the exogenous agent is an inhibitor of retinoic acid synthesis. 
     
     
       15. The composition of  claim 1 , wherein the exogenous agent is selected from the group consisting of BMS-189453, AGN194310, AGN193109, Ro41-5253, SR11335, 9-cis-retinoic acid, disulfiram, and citral. 
     
     
       16. The composition of  claim 1 , wherein the exogenous agent is BMS-189453. 
     
     
       17. The composition of  claim 1 , further comprising a BMP antagonist. 
     
     
       18. The composition of  claim 17 , wherein the BMP antagonist is selected from the group consisting of a BMP 4 antagonist, Noggin, Chordin, Tsg, a BMP soluble receptor, BMPRIA, BMPRIB, and Dorsomorphin. 
     
     
       19. The composition of  claim 1 , further comprising a Wnt inhibitor. 
     
     
       20. The composition of  claim 19 , wherein the Wnt inhibitor is dickkopf homolog 1 (DKK1). 
     
     
       21. The composition of  claim 1 , further comprising a BMP antagonist and a Wnt inhibitor. 
     
     
       22. The composition of  claim 21 , wherein the exogenous agent is BMS-189453, the BMP antagonist is Noggin, and the Wnt inhibitor is dickkopf homolog 1 (DKK1). 
     
     
       23. The composition of  claim 1 , further comprising a culture medium with reduced or depleted vitamin A. 
     
     
       24. The composition of  claim 1 , wherein the mesodermal cell is prepared in vitro by culturing said mesodermal cell on a cell culture plate.